Lin Mingzhang, Piao Linghua, Zhao Zhendong, Liao Li, Wang Dayong, Zhang Haiwen, Liu Xiande
School of Life and Health Sciences, Hainan Province Key Laboratory of One Health, Collaborative Innovation Center of One Health, Hainan University, No. 58 Renmin Avenue, Haikou 570228, China.
Key Laboratory of Tropical Translational Medicine of Ministry of Education & Key Laboratory of Brain Science Research Transformation in Tropical Environment of Hainan Province, School of Basic Medicine and Life Science, Hainan Medical University, Haikou 570228, China.
Pharmaceuticals (Basel). 2025 Jan 9;18(1):67. doi: 10.3390/ph18010067.
Inflammatory bowel disease (IBD) is a persistent inflammatory condition affecting the gastrointestinal tract, distinguished by the impairment of the intestinal epithelial barrier, dysregulation of the gut microbiota, and abnormal immune responses. (L.) , traditionally used in Chinese herbal medicine for gastrointestinal issues such as bleeding and dysentery, has garnered attention for its potential therapeutic benefits. However, its effects on IBD remain largely unexplored. In this study, the major compounds from leaf extract (CCLE) were initially characterized by LCMS-IT-TOF. The IBD model was developed in C57BL/6 mice by administering continuous 4% (/) dextran sodium sulfate (DSS) aqueous solution over a period of seven days. The body weight, colon length, disease activity index (DAI), and histopathological examination using hematoxylin and eosin (H&E) staining were performed in the IBD model. The levels of the main inflammatory factors, specifically TNF-α, IL-1β, IL-6, and myeloperoxidase (MPO), were quantified by employing enzyme-linked immunosorbent assay (ELISA) kits. Additionally, the levels of tight junction proteins (ZO-1, Occludin) and oxidative stress enzymes (iNOS, SOD1, CAT) were investigated by qPCR. Subsequently, flow cytometry was employed to analyze the populations of various immune cells within the spleen, thereby assessing the impact of the CCLE on the systemic immune homeostasis of IBD mice. Finally, 16S rDNA sequencing was conducted to examine the composition and relative abundance of gut microbiota across different experimental groups. In addition, molecular docking analysis was performed to assess the interaction between the principal components of CCLE and the aryl hydrocarbon receptor (AHR). We identified seven bioactive compounds in CCLE: catechin, cajachalcone, 2-hydroxy-4-methoxy-6-(2-phenylcinyl)-benzoic acid, longistylin A, longistylin C, pinostrobin, amorfrutin A, and cajaninstilbene acid. Our results demonstrated that oral administration of CCLE significantly alleviates gastrointestinal symptoms in DSS-induced IBD mice by modulating the balance of gut-derived pro- and anti-inflammatory cytokines. This modulation is associated with a functional correction in M1/M2 macrophage polarization and the Th17/Treg cell balance in splenic immune cells, as well as shifts in the populations of harmful bacteria ( and ) and beneficial bacteria (, unidentified , , and ) in the gut. Furthermore, cajaninstilbene acid, longistylin A, and longistylin C were identified as potential AhR agonists. The present results suggested that CCLE, comprising stilbenes like cajaninstilbene acid, longistylin A, and longistylin C, protects the epithelial barrier's structure and function against DSS-induced acute IBD by restoring gut microbiota balance and systemic immune response as AhR agonists. Overall, CCLE represents a promising natural product-based therapeutic strategy for treating IBD by restoring gut microbiota balance and modulating systemic immune responses.
炎症性肠病（IBD）是一种影响胃肠道的持续性炎症疾病，其特征为肠道上皮屏障受损、肠道微生物群失调以及免疫反应异常。（L.）传统上用于治疗诸如出血和痢疾等胃肠道问题的中药，因其潜在的治疗益处而受到关注。然而，其对IBD的影响在很大程度上仍未得到探索。在本研究中，首先通过液相色谱-质谱联用-离子阱-飞行时间质谱（LCMS-IT-TOF）对（L.）叶提取物（CCLE）的主要化合物进行了表征。通过连续7天给予4%（/）葡聚糖硫酸钠（DSS）水溶液在C57BL/6小鼠中建立IBD模型。在IBD模型中进行了体重、结肠长度、疾病活动指数（DAI）以及使用苏木精和伊红（H&E）染色的组织病理学检查。通过酶联免疫吸附测定（ELISA）试剂盒对主要炎症因子，特别是肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）和髓过氧化物酶（MPO）的水平进行了定量。此外，通过实时定量聚合酶链反应（qPCR）研究了紧密连接蛋白（闭合蛋白（ZO-1）、封闭蛋白（Occludin））和氧化应激酶（诱导型一氧化氮合酶（iNOS）、超氧化物歧化酶1（SOD1）、过氧化氢酶（CAT））的水平。随后，采用流式细胞术分析脾脏内各种免疫细胞的群体，从而评估CCLE对IBD小鼠全身免疫稳态的影响。最后，进行16S核糖体DNA（rDNA）测序以检查不同实验组中肠道微生物群的组成和相对丰度。此外，进行了分子对接分析以评估CCLE的主要成分与芳烃受体（AHR）之间的相互作用。我们在CCLE中鉴定出七种生物活性化合物：儿茶素、卡亚查尔酮、2-羟基-4-甲氧基-6-（2-苯基肉桂基）-苯甲酸、长穗花杉双黄酮A、长穗花杉双黄酮C、豆甾醇、异黄腐醇A和木豆异黄酮酸。我们的结果表明，口服CCLE通过调节肠道来源的促炎和抗炎细胞因子的平衡，显著减轻DSS诱导的IBD小鼠的胃肠道症状。这种调节与脾脏免疫细胞中M1/M2巨噬细胞极化和Th17/Treg细胞平衡的功能纠正以及肠道中有害细菌（和）和有益细菌（、未鉴定的、、和）群体的变化有关。此外，木豆异黄酮酸、长穗花杉双黄酮A和长穗花杉双黄酮C被鉴定为潜在的芳烃受体激动剂。目前的结果表明，包含木豆异黄酮酸、长穗花杉双黄酮A和长穗花杉双黄酮C等芪类化合物的CCLE作为芳烃受体激动剂，通过恢复肠道微生物群平衡和全身免疫反应，保护上皮屏障的结构和功能免受DSS诱导的急性IBD的影响。总体而言，CCLE代表了一种有前途的基于天然产物的治疗策略，可通过恢复肠道微生物群平衡和调节全身免疫反应来治疗IBD。
